A Monocarboxylate Permease of Rhizobium leguminosarum Is the First Member of a New Subfamily of Transporters

Abstract

Amino acid transport by Rhizobium leguminosarum is dominated by two ABC transporters, the general amino acid permease (Aap) and the branched-chain amino acid permease (Bra). However, mutation of these transporters does not prevent this organism from utilizing alanine for growth. An R. leguminosarum permease (MctP) has been identified which is required for optimal growth on alanine as a sole carbon and nitrogen source. Characterization of MctP confirmed that it transports alanine (K m ‫؍‬ 0.56 mM) and other monocarboxylates such as lactate and pyruvate (K m ‫؍‬ 4.4 and 3.8 M, respectively). Uptake inhibition studies indicate that propionate, butyrate, ␣-hydroxybutyrate, and acetate are also transported by MctP, with the apparent affinity for solutes demonstrating a preference for C 3 -monocarboxylates. MctP has significant sequence similarity to members of the sodium/solute symporter family. However, sequence comparisons suggest that it is the first characterized permease of a new subfamily of transporters. While transport via MctP was inhibited by CCCP, it was not apparently affected by the concentration of sodium. In contrast, glutamate uptake in R. leguminosarum by the Escherichia coli GltS system did require sodium, which suggests that MctP may be proton coupled. Uncharacterized members of this new subfamily have been identified in a broad taxonomic range of species, including proteobacteria of the ␤-subdivision, gram-positive bacteria, and archaea. A two-component sensor-regulator (MctSR), encoded by genes adjacent to mctP, is required for activation of mctP expression.Rhizobium leguminosarum is a member of a group of ␣-subdivision of the proteobacteria (the rhizobia), which form a species-specific symbiotic relationship with leguminous plants in order to fix atmospheric nitrogen. The plant supplies the bacteroid (symbiotic bacteria) with a carbon source (C 4 -dicarboxylic acid), while the plant receives reduced nitrogen from the bacteroid. Other rhizobia include Rhizobium, Mesorhizobium, Sinorhizobium, Bradyrhizobium, and Azorhizobium species (8,33,53). Prior to establishing a Rhizobium-legume symbiosis, bacteria must thrive in the soil environment, competing with many organisms for nutrients. Transporters of key nutrients, such as amino acids, may give a competitive advantage to rhizobia, allowing them to better colonize roots. This may account for the large number of transporters that have been revealed by the completed genome sequences of Sinorhizobium meliloti, Mesorhizobium loti, and Agrobacterium tumefaciens (13,26,59).Amino acid transport by R. leguminosarum is dominated by two permeases of the ABC transporter superfamily, the general amino acid permease (Aap) and the branched-chain amino acid permease (Bra). Aap and Bra are members of the polar amino acid transporter (transport classification [T.C.] number 3.A.1.3) and hydrophobic amino acid transporter (T.C. number 3.A.1.4) families, respectively (21, 44). However, both are atypical as they transport a broad range of amino acids (19,56). Indeed, Br...

The Gene yjcG , Cotranscribed with the Gene acs , Encodes an Acetate Permease in Escherichia coli

“…It has also been reported that this protonophore can indirectly dissipate the sodium concentration gradient depending on the activity of Na ϩ /H ϩ antiporters. This may result in the inhibition of Na ϩ -dependent transport systems, as has been reported for proline transport (31) or sodium-dependent GltS glutamate transport, both in E. coli (10). Our results showed that the two entry processes are sensitive to CCCP (Fig.…”

“…The specificity of ActP is rather narrow, a common feature of most bacterial carboxylate permeases, such as GlcA and LldP; an exception is the monocarboxylate permease of Rhizobium leguminosarum, which displays wider specificity (10). In this context, it is of interest that although ActP, like the GlcA and LldP permeases, transports small carboxylates, there is no sequence similarity between these transporters and ActP.…”

“…It is known that it is difficult to completely deplete sodium in the reaction mixtures and buffers used in transport assays, especially under our conditions, under which sodium was present in the labeled source of acetate. The residual sodium levels might be sufficient to allow transport by systems with a high affinity for this cation (10). In this context, the inhibition of acetate transport by the uncoupler CCCP indicates that the driving force used by ActP is a transmembrane electrochemical potential.…”

Systems approach to refining genome annotation

“…Although IdnT is a known D-gluconate and L-idonate transporter, a prior study has shown that 5-keto-D-gluconate inhibits D-gluconate transport (15). PutP is a proline transporter that belongs to the same family of transporters as a propionate transporter (MctP) from Rhizobium leguminosarum (16), and the deletion of putP reduces the growth rate ( ) on propionate ( ϭ 0.02 Ϯ 0.002 h Ϫ1 for ⌬putP as compared with 0.06 Ϯ 0.002 h Ϫ1 for the parental strain). This combined evidence suggests that putP and idnT are responsible for the transport of propionate and 5-keto-Dgluconate, respectively.…”

Variation among Desulfovibrio Species in Electron Transfer Systems Used for Syntrophic Growth

“…However, both lactate transporters might be required to drive the flux of lactate needed to support syntrophic growth. Alternatively, LctP-2 could be a misannotated acetate permease, since the sequence identity to the known lactate permeases is low (Ͼ30%) and homologs to acetate transporters of bacteria (e.g., ActP in E. coli [38,39]) are missing in the D. alaskensis strain G20 genome.…”